NEUROPROTECTIVE EFFECT OF PIPER CUBEBA L.F. (PIPERACEAE) FRUIT EXTRACT AND NANOEMULSION IN ISCHEMIC STROKE ANIMAL MODEL

• Main Author: Putri Dwita, Lusi
• Format: Dissertations
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/75995
• Institution: Institut Teknologi Bandung

Neuroprotection is an effort to protect neurons to reduce nerve damage in conditions such as Parkinson's, traumatic brain injury and ischemic stroke. Several mechanisms of natural products that have been known to act as neuroprotectors are through the suppression of oxidative stress and neuroinflammation, which are the main causes of neurological disorders. The plant suspected of having the potential to have this effect is Piper cubeba L. f., known in Indonesia as kemukus. This study aimed to find potential candidates for neuroprotective agents that can be used in neurological disorders, one of which is ischemic stroke. The study began with the preparation of test materials consisting of 70% ethanol extract (PC70), 96% ethanol extract (PC96), lignan fraction (DCM) and essential oil (PO) of P. cubeba fruit. Next, the characterization of the extract was carried out. Analysis of the chemical compound content of the extract was carried out by thin layer chromatography (TLC), liquid chromatography–mass spectrometry (LC-MS) and gas chromatography-mass Spectrometry (GC-MS). Pharmacological testing began with testing the antioxidant activity in rat brains, and then the test material with the best activity was continued with neuroprotective tests in animals with cognitive impairment. The best results from this test were then formulated into a nanoemulsion, after which it was tested on the antioxidant activity of the P.cubeba nanoemulsion in the brain with two routes of administration, namely orally and intranasally. Finally, the neuroprotective activity of P. cubeba nanoemulsion was tested in ischemic stroke animals model. In the initial study, the results of the antioxidant effect test on rat brains showed that all test groups (PC96, PC70, DCM and PO) had the potential to significantly (p<0.05) inhibit lipid peroxidase compared to controls. CAT activity in all groups showed a significant (p<0.05) increase compared to control, except in the PC70 group. All test groups also showed significantly (p<0.05) lower NO levels compared to the control group. Based on the results of this study, it can be concluded that P. cubeba has the potential as a neuroprotector with brain antioxidant activity, especially in the PC96 and DCM groups. The second test was conducted on an animal model with cognitive impairment using Electroconvulsive Shock (ECS). Animals' cognitive function was tested with Morris water maze (MWM), and the brains were taken for lipid peroxidase, SOD, CAT, TNF-? and IL-1? tests. The results of the MWM test showed that PC96 and DCM improved cognitive function significantly (p<0.05), marked by a decrease in escape time of up to 70% better than the ECS control. The results of brain homogenate examination showed that PC96 and DCM significantly (p<0.05) increased CAT activity in the hypothalamus and cerebral cortex compared to ECS controls. While the increase in SOD activity only occurs in the cortex. The DCM group only significantly (p<0.05) increased CAT activity in the hippocampus and SOD in the cortex compared to ECS controls but did not affect lipid peroxidase. Administration of PC96 and DCM in the ECS model also significantly (p<0.05) reduced TNF-? and IL-1? levels compared to the ECS control and was comparable to citicoline. Then PC96 and DCM are formulated into nanoemulsions. The results of the thermodynamic stability test of the nanoemulsion extract (NE) and the (NF) fraction of P. cubeba showed that the nanoemulsion was stable, and there was no significant change in size or polydispersity index (PDI). NE and NF were added with 0.5% chitosan for intranasal administration (KNE and KNF). Brain’s catalase activity test results showed NE (p.o), NF (p.o), intranasal KNE and intranasal NE, significantly (p<0.05) increased catalase activity in the hippocampus compared to controls. However, only NE (p.o) and intranasal KNE significantly (p<0.05) increased catalase activity in the prefrontal cortex compared to controls. Nanoemulsion (PCN) and 96% ethanol extract of P. cubeba (PCE), continued in neuroprotective test in ischemic stroke animal model. Ischemic stroke were induced using the tMCAO model. The results showed that tMCAO induction with 1-hour occlusion caused mild to moderate neurological disorders. Giving PCN and PCE reduced the severity of neurological disorders to mild disorders. The results of brain staining with TTC showed significant (p<0.001) lower infarction in the PCN group compared to MCAO control group. The results of the brain homogenate test showed that PCN significantly (p<0.001) inhibited lipid peroxidase compared to the MCAO control, and increased catalase and glutathione activity significantly (p<0.001) compared to the MCAO control. Likewise, with inflammatory cytokines in the brain, PCN significantly (p<0.05) reduced TNF-? concentrations, especially in the infarcted hemisphere, compared to MCAO controls. However, the decrease in IL-1? was better in the PCE group. Only PCN100 showed a significantly (p<0.05) increased anti-inflammatory cytokine (IL-10) compared to the MCAO control. Therefore, it can be concluded that both PCN and PCE showed neuroprotective activity in tMCAO-induced animals, with the best activity shown by PCN. Based on the study's overall results, it can be concluded that P. cubeba in the form of nanoemulsion had more potential as a neuroprotector than the extracts and can be developed as a neuroprotective agent that reduced the impact of ischemic stroke reperfusion injury by increasing antioxidants, decreasing proinflammatory cytokines and increasing antiinflammatory cytokines in the brain.